The goal of the proposed Phase I research is to develop a set of commercial reagents for recognizing 2'-O-methyladenosine (Am), 2'-O-methylcytidine (Cm), 2'-O-methylguanosine (Gm) or 2'-O-methyluridine (Um), as well as indiscriminately to all four 2'-O-methyl ribose moiety. The core technologies that serve as the foundation of these new reagents are based on the unique properties of camelid antibodies, which have been well accepted as valuable binding reagents for proteins and haptens, but so far without any example for RNA modifications. The main advantage of the proposed single-domain (VHH) antibodies from camelidae is that they have very high affinity and specificity, and they are easy to reproduce as monoclonal antibodies, stable, and of very small size, enabling super-resolution imaging when conjugated to dye. Currently, affinity reagents of any type are only available for monitoring a handful among the more than 100 RNA modification moieties. The best examples of such available reagents are the antibodies against N6-methyladenosine (m6A), which proved to be of significant value to researchers in the field trying to understand the functions of the m6A-modified mRNAs. However, the two anti-m6A antibodies, created about 3 decades ago, are both rabbit polyclonal antibodies that are not easily reproducible or suitable for high precision immune-imaging or other modern assays. There is an urgent need for development of high quality antibodies against the fast-expanding and increasingly important family of known modified mRNAs and regulatory RNAs. Successful execution of the proposed project will establish a fully tested, market-ready group of reagents and kits for detecting one type of closely related RNA modifications, 2'-O-methylation on the ribose of the 4 RNA bases. Currently, there is a lack of such reagents on the market, providing the applicant a very good chance of commercial success with the outcome of the proposed project. After achieving this goal, similar efforts can be initiated in a phase II SBIR or other types of project to develop more nano-antibodies against additional RNA modifications, eventually overcome the bottleneck of post-transcriptional RNA studies, and serve as the foundation for advancing NIDA- and NIMH-relevant research projects on RNA influence in various substance abuse or mental diseases.